Effects of lysine modification on the structural and physicochemical properties of melanin from Morchella importuna

Melanin is a natural ingredient produced widely in edible fungi with excellent bioactivity. The edible fungi can provide a rich source of melanin. Morchella importuna (MIM), a precious edible and medicinal fungus, has been domesticated in China in recent years. However, the extraction of melanin from MIM has been rarely reported. Besides, the hydrophobicity of melanin can limit its applications. Hence, it is a high demand to efficiently extract and improve the water solubility by modification. Alkali-extraction and acid-precipitation with ultrasound-assisted approach can often be used to extract melanin. Furthermore, it has not yet been reported to extract melanin from MIM using cellulase and ultrasonic. In cellulase extraction, the cellulase can be used to degrade cellulose components in the microbial cell wall, in order to increase the extraction rate. In ultrasonic extraction, ultrasonic vibration can be used to make the microbial tissue structure form a cavity combine with its thermal effect. The better extraction of melanin can be obtained, in terms of the short extraction time and high extraction rate. It is expected to combine the cellulase and ultrasonic extraction for the active ingredients of natural products. In this study, the extraction process was optimized by single-factor, Plackett-Burman and response surface test, in order to improve the extraction rate of the melanin from MIM. The solubility of MIM was improved by modification with lysine. The structural and physicochemical properties of MIM were evaluated before and after modification, together with solubility and stability, using UV-visible absorption, FTIR spectra, scanning electronic microscope (SEM). The results showed that the highest extraction rate of MIM was achieved under the conditions of NaOH solution concentration of 1.54 mol/L, cellulase addition of 20 mg/g, cellulase digestion time of 78.6 min, material-liquid ratio of 1:30, digestion temperature of 40 °C, and ultrasound time of 80 min. The absorbance of modification was compared with four amino acids, in order to determine the optimal amino acid and mass ratio. The water solubility was significantly improved after lysine modification. The color values of MIM and lysine-modified Morchella importuna melanin (L-MIM) were 480.24 and 1 771.18, respectively. The solubility and color value of L-MIM were higher than those of MIM. The maximum absorption peaks of MIM and L-MIM were 230 and 219 nm, respectively. The absorption of MIM and L-MIM showed a decreasing trend with the increasing wavelength. It was also found that the MIM and L- MIM had no special absorption peaks at 260 and 280 nm, indicating less nucleic acids and proteins without impurity peaks. The FTIR spectra demonstrated the typical indole structure of melanin. SEM tests revealed that there were the smooth surface and uniform cubic crystal structure in MIM samples, whereas, the L-MIM samples were in the form of large lumps, with the raised surfaces that adhered to each other. The reason was that the particle adhesion was susceptible to moisture absorption in the L-MIM samples. In addition, both MIM and L-MIM were relatively stable under the temperature from 20 to 100 ℃, in the dark and natural light, while both were basically stable, when pH value from 8.0 to 12.0. It was found that Fe³⁺ had a significant impact on both MIM and L-MIM stability. These findings can provide the clear directions for their applications in functional foods.